In search of silver bullets

George Vassiliou on his work in the battle against Acute Myeloid Leukaemia

The outlook is bleak - four out of every five patients with Acute Myeloid Leukaemia die from their disease. This
dreadful prognosis for his patients has driven clinician scientist Dr George Vassiliou to join the search for genetic
changes underlying the disease in the quest to improve the treatment, survival and quality of life of sufferers.

It has been estimated that, each day, 500 billion of our cells divide. One of the remarkable features of our
multi-cellular, human life is that cell division so rarely goes badly wrong.

Like all complex creatures, we owe our existence to the complicated, coherent, choreographed cooperation among the
cells that make us up. We live because they live and we thrive because they perform. We grow and our tissues repair
themselves because our cells divide when needed. But being a multi-celled creature brings us dangers: if our cells
divide when not required, if the checks and balances that keep everything in order fail, we face cancer.

Cancer is a near inevitable part of our complex life, often arising from mutations that result in loss of control of
cell growth and division. The treatment of cancer traditionally relies on the use of chemotherapy and radiotherapy;
potent treatments that can destroy cancer cells but usually also harm healthy cells, leading to unpleasant side
effects. However, in the last two decades, researchers have uncovered some of the genetic changes that underlie many
cancers and such discoveries are accelerating dramatically with the advent of increasingly sophisticated scientific
technologies.

As a result, many are hopeful that these discoveries will be harnessed to improve the treatment of cancer by
targeting the specific mutations involved in its development. In practice, a series of steps need to be taken to
develop targeted anti-cancer treatments: the first is to classify cancers according to the types of cells they affect
and their clinical behaviour; the second step is to identify and validate the genes responsible for each type of
cancer as appropriate drug targets; the final step is to develop drugs against the targets and to show them to be
clinically effective and safe to use in patients. This process is often referred to as 'rational drug design'.

"Traditional anti-cancer chemotherapy and radiotherapy often give rise to unpleasant toxicities
due to harmful effects on normal cells," says George Vassiliou, a clinician scientist at the Sanger Institute.
"Moreover, the effectiveness of such therapies is often modest or short-lived. However, the era
of 'rational drug design' is ushering in new hope that we will be able to develop effective therapies with fewer
side-effects."

George is both a researcher at the Sanger Institute and a clinician at Addenbrooke's Hospital in Cambridge, and is on
the hunt for 'rationally' designed drug treatments for Acute Myeloid Leukaemia (AML). AML represents an uncontrolled
production of highly abnormal blood cells in the bone marrow, which if left untreated leads to the rapid demise of
the patient. The cell proliferation causes clogging of the bone marrow and prevents the production of normal blood
cells. Doctors can sometimes treat AML successfully using chemotherapy, radiotherapy or bone marrow transplantation,
but the long-term survival rates remain disappointingly low. It is by looking at the genetic basis of AML, that
George hopes to assist in the development of molecular treatments that will be of real and lasting benefit to his
patients.

"By applying our molecular understanding of cancers such as Acute Myeloid Leukaemia (AML), we can
hope to develop 'intelligent' drugs that target the disease whilst sparing normal cells," says George. "Clinically,
this should translate into improved survival and quality of life for patients."

The revolutionary drug Glivec® was the first example of a drug developed rationally using the findings of
molecular research. It was developed to treat Chronic Myeloid Leukaemia (CML) in the late 1990s and probably
represents the single most significant development in the history of cancer treatment.

"We need a Glivec for Acute Myeloid Leukaemia," says George.

The Glivec® Story

Chronic Myeloid Leukaemia (CML) is a myeloproliferative disorder, a disorder that is characterised by a massive and
uncontrolled expansion of myeloid cells in the bone marrow and blood. Unlike AML, which affects around 2,300 people
in the UK each year and can make sufferers very ill quickly, CML affects approximately 750 people in the UK each
year, and can remain clinically stable for many years.

CML is associated with the presence of the Philadelphia chromosome, an abnormal chromosome composed of parts of
chromosomes: 9 and 22. This creates a fusion of part of the BCR gene from chromosome 22 to part of the ABL gene on
chromosome 9. The BCR-ABL fusion gene produces an abnormal tyrosine kinase protein that is always active or 'switched
on'. The leukaemia develops as a result of uncontrolled cell growth triggered by the effects of this protein on cell
growth and proliferation.

Glivec was identified in the late 1990s by a team of chemists from the Swiss pharmaceutical company Ciba-Geigy, now
Novartis, during a targeted screen for tyrosine kinase inhibitors. It was found to specifically inhibit the
proliferation of blood cells with the BCR-ABL fusion, and was later developed clinically in collaboration with Dr
Brian Druker, from Oregon Health and Science University.

Glivec is one of the most striking developments in cancer treatment since medicine began. It has laid the path for
the development of other drugs that work against specific gene targets.

George's research

Normal blood cells (top) and blood from a patient with AML (bottom). The AML sample contains a large number of abnormal (leukaemic) cells.

George's novel molecular approach to the study of AML aims to identify new drug targets and ultimately to improve the
treatment of patients. The clinical onset of AML can take place at an alarming rate, sometimes taking hold in just
days or weeks. Even with the best available therapies, the survival rate is low.

The most common genetic mutation in AML affects a gene called NPM1, and this gene is in the centre of George's work.
In simple terms George wants to answer two questions: first, how does the mutation in NPM1 promote the development of
AML? And, second, what other genes work together with NPM1 in this process?

"If we answer these questions, we will have made a significant forward step towards the design of
improved treatments for AML patients," explains George. "That's why we have developed and
are currently studying a mouse model of AML carrying this mutation."

The mouse model, developed at the Sanger Institute, carries a silent version of the NPM1 mutation found in human AML
patients. This silent version has no effect on the mouse. By treating these mice with a drug, researchers can 'switch
on' a protein called recombinase, which activates the mutation in the blood cells of mice. The mice are then studied
to identify the effects of the mutation on blood development and its role in the development of leukaemia.

But mutations in NPM1 are only one part of a far more complex story; the mutation in the NPM1 gene is thought to
cooperate with mutations in other genes to cause AML. To try to tease out this complex set of interactions between
mutations, these same mice will also be studied using a technique called 'insertional mutagenesis'. This specialised
technique allows researchers to accelerate the development of cancers by randomly altering genes, whilst at the same
time 'tagging' the altered genes thus making them easy to identify. Only a handful of research institutes have the
expertise to carry out insertional mutagenesis, but it is this crucial technique that will facilitate the discovery
of genes that co-operate with NPM1 to cause AML.

"Insertional mutagenesis is being used in mice carrying the human leukaemia-causing mutation in
NPM1," explains George. "We hope that this will elucidate its cancerous effects and help
to identify other co-operating genes involved in the development of AML."

Why use mice?

In cancer research, it is not always possible to extrapolate results from studies performed in the test tube - in
vitro - to effects on whole organism. Drugs that are effective on cell lines in culture might cause unexpected
toxicity in patients, or they could simply be ineffective. In the case of NPM1, it is possible only to draw limited
inferences from in vitro studies. Studies in living organisms - in vivo - can provide far more information about the
role of NPM1 in the development of AML and the identity of new treatment targets.

Mice are an important and widely used animal model for the study of human development and disease. Mouse models can
be generated through the targeted manipulation of mouse embryonic stem cells to generate many identical mice carrying
genetic mutations associated with human disease. The technology, that made this possible, earned its pioneers -
Martin Evans, Mario Cappechi and Oliver Smithies - the 2007 Nobel Prize for Medicine or
Physiology, and is now in widespread use in biomedicine.

"This research is producing tantalising results," says George. "We are
beginning to understand the effect of NPM1 on blood development, we recently observed the first mice with leukaemia
and we have the first clues about genes working in concert with NPM1 to cause leukaemia. But these exciting
developments are just the first steps on the road towards a detailed understanding of the role of these genes in
leukaemia."

At the Sanger Institute, George works closely with his colleagues in Mouse Genomics, the sequencing teams, the
microarray facility, and the Cancer Genome Project. He also collaborates with the Dr Derek Stemple's group, who work
with another model organism, the zebrafish; as well as Dr David Adams and Dr Pentao Liu, whose groups work in
experimental cancer genetics.

His story

Localisation of normal (top) and mutant (bottom) NPM1. The mutant loses its discrete location in the nucleolus.

George studied medicine in London where he later started his training as a haematologist. He then went on to study
for his PhD in myeloproliferative disorders, in Professor Tony Green's laboratory at the Department of Haematology,
University of Cambridge. After becoming a Haematology specialist, he had the choice of becoming a clinical consultant
in the NHS, or to follow a career in medical research. He chose research and approached Professor Allan Bradley, the
Sanger Institute's director, with his ideas for studying AML.

"I was thoroughly impressed by the ideas George brought with him to the Sanger Institute,"
says Allan Bradley. "He was convinced that by looking at the complex of interactions and
processes that underlie Acute Myeloid Leukaemia, researchers had the potential to find new drugs to target this
cancer. All of his research ambitions are driven by his determination to improve the prognosis of his
patients.

"The Sanger Institute strives to make a difference to understanding of biology and ultimately to
healthcare and clinical appointments, such as George's, are an important part of our contribution."

With Allan's support, in 2006 George applied for and was awarded a five-year fellowship from Cancer Research UK to
fund his position and his research.

"My time at the Sanger has been truly exciting and I've met many remarkable people," he
says. "Allan has been an excellent supervisor, offering sound advice and direction whilst
nurturing intellectual freedom and enterprise. I've also had valuable help and advice from my Cancer Research UK
mentor, Professor Mike Stratton, who directs the Cancer Genome Project at the Sanger Institute."

George strikes a balance between his research at the Sanger Institute, and his role as a clinician. Every Tuesday
morning, he runs an NHS out-patient clinic at Addenbrooke's Hospital, and for one month each year he has day-to-day
charge of in-patients undergoing chemotherapy and bone marrow transplantation for leukaemia and other blood-derived
cancers at the hospital. George also helps to develop the next generation of clinicians, by teaching Cambridge
University medical students, and - unfortunately for them - also examining them. Balancing his research demands, with
his clinical obligations is a challenge, not to mention his young family, who he also likes to see occasionally!
However, he declares unequivocally that he loves what he does and he would not change it for any other job.